Proprotein convertase subtilisn/kexin 9 (PCSK9) is a member of the mammalian PCSK family that, to date, includes eight other members; PCSK1 (PC1/3), PCSK2 (PC2), PCSK (Furin), PCSK4 (PC4), PCSK5 (PC5/6), PCSK6 (Pace4), PCSK7 (PC7) and PCSK8 (SKI-1/SIP) [1]. Collectively this family is responsible for the proteolytic maturation of secretory precursors to bioactive proteins and peptides including neuropeptides, pro-hormones, cytokines, growth factors, receptors, cell surface proteins and serum proteins [2, 3]. Fitting with its role in cholesterol metabolism PCSK9 is highly expressed in the liver and intestine, two tissues important in cholesterol homeostasis [4]. It is also found in circulation [5-7]. PCSK9, like its family members, is synthesized as a preproprotein containing several defined motifs; a signal peptide domain for routing the PCSKs to the secretory pathway, a prodomain important for folding and acting as an endogenous inhibitor, a catalytic domain characteristic of serine proteases, and a C-terminal Cys and His rich domain (CHRD) implicated in enzyme stability and protein-protein interaction [3]. We reported that PCSK9 is autocatalytically processed in the endoplasmic reticulum (ER) at the site FAQ152↓SIP indicative of its consensus cleavage motif, travels to the Golgi where its sugar residues at the glycosylation site N533CS are matured and its propeptide is sulfated at Tyr38, and is secreted [4,5]. PCSK9 is unique among the PCSK family because it is secreted in association with its inhibitory propeptide.
Cell culture and animal models have established that the low density lipoprotein receptor (LDLR) is one of the main down-stream targets of PCSK9 [4, 8-11]. Supporting this, several groups have reported that secreted PCSK9 can interact with and enter the endocytic recycling pathway with LDLR, affecting the equilibrium of LDLR recycling versus LDLR lysosomal-dependent degradation [6, 12-15]. The ‘gain of function’ D374Y variation in the catalytic domain of PCSK9 results in the most severe form of autosomal dominant hypercholesterolemia (ADH) [16,17]. Studies have shown that this variant binds the LDLR receptor (within its epidermal growth factor (EGF)-A domain) at the cell surface 25× more efficiently than wild type PCSK9 thereby shifting the equilibrium toward LDLR lysosomal-dependent degradation [12,15]. However the effect of other ADH-associated PCSK9 variants, such as the PCSK9(S127R) on PCSK9-LDLR dependent degradation is less obvious since their binding equilibrium to the LDLR is only moderately increased [15,18]. Crystal structures have shown that this Ser127 residue does not interact directly with the LDLR [19].
Longitudinal population studies have shown significant reduction in the risk of coronary heart disease (CHD) in ‘loss of function’ PCSK9 carriers [20,21]. Reduced plasma PCSK9 concentrations for at least three PCSK9 variants, R46L, Y142X and C679X increase the amount of LDLR that is recycled, effectively reducing plasma LDL cholesterol (LDLC) [7,22]. As is the case with ‘gain of function’ PCSK9 variants not all ‘loss of function’ variants can be attributed to a single mechanism, in this case, reduced plasma PCSK9. However these studies, along with the identification of two healthy PCSK9 ‘null’ individuals [7,23] have generated much interest toward understanding the exact details of the mechanism(s) of PCSK9-dependent:LDLR degradation, its site(s) of action, whether the effect is direct or indirect, and how different PCSK9 SNPs alter its function. It is believed that the design of PCSK9 inhibitors may provide a promising therapy for treatment of hypercholesterolemia [7, 11, 24].
There is a need in the art for novel compounds that alter the interaction of PCSK9 with LDLR. There is also a need in the art for compounds that either inhibit or enhance LDLR degradation. Further, there is a need in the art for novel compounds that increase the amount of recycled LDLR thereby decreasing plasma low density lipoprotein C (LDLC). There is also a need in the art for novel compounds that alter the normal biological function of PCSK9. Further, there is a need in the art to understand and manipulate the mechanisms by which PCSK9 interacts with specific proteins including LDLR. There is also a need in the art to identify novel compounds and compositions that can modulate normal PCSK9 phosphorylation. Further, there is a need in the art for novel methods that may be employed to modulate PCSK9 phosphorylation.